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  • Dyna, ao 78, Nro. 168, pp. 72-80. Medelln, Agosto, 2011. ISSN 0012-7353

    CHARACTERIZATION OF NATURAL MICROCOSMS OF ESTUARINE MAGNETOTACTIC BACTERIA

    CARACTERIZACIN DE MICROCOSMOS NATURALES DE BACTERIAS MAGNETOTCTICAS ESTUARINAS

    ALEJANDRO SALAZAR Escuela de Biociencias, Universidad Nacional de Colombia, Medelln. asalazar1370@gmail.com.

    ALVARO MORALESGrupo de Estado Slido, Instituto de Fsica, Universidad de Antioquia, A.A. 1226, Medelln, Colombia. amoral@fisica.udea.edu.co

    MARCO MRQUEZFacultad de Minas, Universidad Nacional de Colombia, Medelln. mmarquez@unalmed.edu.co

    Received for review May 24th, 2010; accepted December 3rd, 2010; final version October 27rd, 2010

    ABSTRACT: To date, no complete study of magnetotactic bacterias (MTB) natural microcosms in estuarine or tropical environments has been reported. Besides, almost all the studies around magnetotactic bacteria have been based on fresh waters away from the Equator. In this work, we focused the experimental region at the Equator and present a comprehensive mineralogical and physicochemical characterization of two estuarine bacterial microcosms. The results show that mineral lixiviation in the sediments may be an important factor in the solubilization of elements required by magnetotactic bacteria. Specifically, we show that clinochlore, phlogopite, nontronite, and halloysite could be among the main minerals that lixiviate iron to the estuarine microcosms. We conclude that nitrate concentration in the water should not be as low as those that have been reported for other authors to achieve optimal bacteria growth. It is confirmed that magnetotactic bacteria do not need large amounts of dissolved iron to grow or to synthesize magnetosomes.

    KEY WORDS: Magnetotactic bacteria (MTB), magnetosome, microcosm, estuary

    RESUMEN: No se ha reportado ningn estudio completo sobre microcosmos naturales de bacterias magnetotcticas (MTB) en estuarios o ambientes tropicales. Adems, casi todos los estudios sobre las bacterias magnetotcticas se han desarrollado en aguas dulces alejadas del ecuador. Este trabajo se desarrolla sobre el ecuador y reporta una caracterizacin mineralgica y fisicoqumica detallada de dos microcosmos bacterianos estuarinos. Los resultados muestran que la lixiviacin de minerales en los sedimentos puede ser un factor importante en la solubilizacin de elementos requeridos por las bacterias magnetotcticas. Especficamente, que el clinocloro, flogopita, nontronita y haloisita pueden estar entre los minerales ms importantes en la lixiviacin de hierro a los microcosmos estuarinos. Se concluye que la concentracin de nitrato en el agua no debe ser tan baja como se ha reportado para lograr un crecimiento bacteriano ptimo. Las bacterias magnetotcticas no necesitan grandes cantidades de hierro disuelto para su crecimiento ni para la sntesis de magnetosomas.

    PALABRAS CLAVE: Bacterias magnetotcticas (MTB), magnetosomas, microcosmos, estuario.

    1. INTRODUCTION

    Magnetotactic bacteria are microorganisms of the bacteria domain, whose directional swimming behavior is affected by the Earths geomagnetic and external magnetic fields [1-2]. This property is known as magnetotaxis [3-4]) and occurs mainly due to the presence of magnetic nanocrystals (generally of magnetite [Fe3O4], or gregite [Fe3S4]) that shape an intracellular, single-magnetic-domain and membrane-bounded structure known as magnetosome [2, 4]. This property is generally assumed to facilitate the bacteria in its finding and maintaining a favorable position in

    vertical chemical gradients in stratified environments [5, 6]. Currently, a wide morphology variety of MTB has been reported, such as coccus, bacillus, vibrio, spirillum, and multicellular aggregates [2, 7, 8].

    Many authors have studied the natural environment of different species of MTB, searching for a strategy to obtain large amounts of magnetic nanocrystals [4, 9, 10]. The objective of those studies was to identify the most important physico-chemical factors that are involved in the growth of MTB populations and the synthesis of magnetosomes [1, 4, 10]. Some of these chemically dissolved factors, are the iron (total Fe, Fe2+, and Fe3+), sulfates, nitrates in the solution,

  • Dyna 168, 2011 73

    and dissolved oxygen (DO) [1, 4, 11, 12]. Furthermore, all the reported MTB are anaerobic or strict-microaerophilic [7, 12], and mesophilic [1, 10]. Another factor that could be important in those processes is the microbial ecology of the natural environments of MTB, but it has not yet been studied thoroughly [1].

    In this research, different spectroscopic techniques and chemical analyses were used to characterize two estuarine MTB microcosms, situated in the tropical waters of the Caribbean Sea. We report the variations in soil mineralogy, water composition, and some physico-chemical parameters of the MTB-environments. This information may serve as a guide to elucidate potential mineral donors that contribute to the formation of magnetosomes and other intracellular bodies.

    2. MATERIALS AND METHODS

    2.1 Sampling zone

    The samples were taken in two different estuaries. Cispat Bay (BC) (921 - 925 North latitude, 7545 - 7550 West Longitude) and the Caimanera Bog (CC) (925 north latitude, 7541 east longitude) (Fig. 1). Both estuarine systems are located in the Morrosquillo Gulf in the Colombian Caribbean Sea and are conformed by a great variety of mangrove swamps that shelter an abundant population of marine and estuary species.

    Figure 1. Location of sampling zone: Cispat Bay (BC) and Caimanera Bog (CC) in the Morrosquillo Gulf, Colombia.

    The water-sediment samples were taken in the Oxic-Anoxic Transition Zone (OATZ), in northern regions of both estuaries. A HANNA oxymeter was used to locate the OATZ in the water column. The depth of sampling in BC and CC was around 2 m and 1.5 m, respectively.

    2.2 Spectroscopy

    The spectroscopy techniques used to study the composition of the sediments were: X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and Mssbauer spectroscopy (MS). Before the analyses, the sediments were dried at room temperature and milled to pass through a 200 Tyler mesh.

    2.2.1 X-ray Diffraction

    The XRD analyses were carried out in Panalytical Xpert Pro MDP equipment, in a 2 range of 10 to 70, at a speed of 0.02/s, using Cu k radiation with a current of 40 mA and 45 kV. The results were analyzed with the diffraction software DIFFRACplus 2000 and the data base PDF2.MDI.

    Relative abundance of the minerals in the sediments was qualitatively estimated based on the height of the peaks in the spectra.

    2.2.2 Fourier transform infrared spectroscopy

    The FTIR analyses were carried out in a Spectrum One Perkin Elmer Spectrophotometer, operating by transmittance between 4000 and 450 cm-1. The pellets were made with milled sediments and KBr.

    2.2.3 Mssbauer spectroscopy

    Wissel Mssbauer equipment was used in transmission and constant acceleration mode for Mssbauer spectra (MS) acquisition. The equipment was operated with 57Co in a rhodium matrix.

    2.3 Water analysis and physico-chemical parameters

    2.3.1 Water analysis

    The codes of the methods used for the water analyses refer to the STANDARD METHODS FOR EXAMINATION OF WATER AND WASTEWATER [13]. Total alkalinity (mg/L CaCO3) (2320B); alkalinity to phenolphthalein (mg/L CaCO3) (2320B); chlorides (mg/L Cl

    -) (4500-CI B); phosphates (mg/L PO4

    3--P) (4500-P D); total phosphorus (mg/L P) (4500-P D); nitrates (mg/L N-NO3

    -) (4500- NO3- A); nitrites (mg/L N-NO2

    -) (4500- NO2-B); ammoniacal nitrogen (mg/L NH3-N) (4500- NH3 C.D); total nitrogen

  • Salazar et al74

    (mg/L N) (4500 - NORG); organic nitrogen (mg/L N) (4500 - NORG); total, ferrous and ferric iron (3500 Fe); sulfates (4500 SO4

    2-).

    2.3.2 Physico-chemical parameters

    We used a digital thermometer, HACH HQ40d pH-meter, SCHOOTT Eh-meter and HI 9143 HANNA oxymeter, to measure temperature, pH, Eh, and dissolved oxygen (DO), respectively. All the measurements were in situ. Average salinity was calculated in the laboratory by the evaporation of a fixed volume of estuary water. Salinity was calculated as an average.

    2.4 MTB and magnetite nanocrystals presence

    MTB from BC and CC were magnetically isolated using the glass recipient described by Ulysses Lins et al. (2003) [14]. Magnetotaxis were confirmed by optical microscopy (Olympus CX31). Magnetite nanocrystals were detected by electronic microscopy (Phillips, Tecnai G2), operating at 200kV, and identified by Energy-dispersive X-ray spectroscopy (EDX). Additionally, MTB population was counted in natural BC and CC samples using a BOECO Neubauer chamber.

    3. RESULTS

    3.1 X-ray Diffraction

    Figure 2 shows the XRD spectra for BC and CC sediments. Quartz appears as the most abundant mineral in both sediments. No magnetite or gregite were detected by XRD in BC or in CC. However, MTB (and its magnetite magnetosomes) presence in the systems was confirmed by optical and electronic microscopy.

    Figure 2. XRD spectrum of a sediment sample from BC

    (up) and CC (down). The graphics are on d-scale.

    3.2 Fourier transform infrared spectroscopy

    Figure 3 shows the FTIR spectra of BC and CC.

    Figure 3. FTIR spectrum of a sediment sample from BC (up) and CC (down).

    Bands in 3420 cm-1 and 1645 cm-1 (Fig. 3, left), and in 3441 cm-1 and 1645 cm-1 (Fig. 3, right), are commonly associated with vibrations of the H-O bonds in water molecules [15, 16]. Bands in 2928 cm-1 and 2862 cm-1 are reported as vibrations of C-H bonds in organic compounds [16]. The little band in 2357 cm-1 is associated with vibrations in the CO2 molecule [15, 17]. Bands on the right

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